Abstract: Necrotizing enterocolitis (NEC) is a leading cause of morbidity and mortality in the neonatal intensive care unit (NICU). Because current therapies remain unsatisfying and often unsuccessful, research on the development of preventive strategies is now a stated priority. The pathogenesis of NEC is poorly understood. A leading hypothesis in the field is that an injury sustained during the neonatal period elicits an exaggerated inflammatory response by the immature intestinal epithelial cells of the premature infant. Furthermore, recently, aberrant or excessive apoptosis of the gut epithelia has been recognized as either an initiating or necessary step in the pathogenesis of NEC, and abnormal bacterial colonization of the premature infant intestine may exacerbate the pathological process. Recent laboratory and clinical studies suggest that NEC may be prevented by probiotics, which may normalize bacterial populations within the gut of premature infants and also promote cytoprotective intestinal responses. Probiotic commensal bacteria can affect intestinal epithelial responses through TLR signaling, which can regulate intestinal epithelial host defenses by improving cell survival and by mitigating inflammatory responses to injury. Furthermore, commensal bacteria can also generate low levels of reactive oxygen species (ROS), which activate proliferative signaling and prevent inflammatory signaling in the intestinal epithelial cell by inactivating the specific ubiquitin ligase complex responsible for -catenin degradation and NF-B activation respectively. We hypothesize that probiotics may help in the prevention of NEC not only by normalizing gut flora but also by directly improving host defense through improved cell survival and reduced proinflammatory mediators, via TLR and ROS signal transduction. Our long term goal is to explain how the probiotic commensal bacterium Lactobacillus rhamnosus (LGG) might act to prevent NEC in newborn humans. In this proposal, we aim to explain how the probiotic commensal, LGG reduces apoptosis, promotes proliferation, and inhibits excessive inflammation in the developing intestine of the premature infant using a murine model of immature intestines. Future studies with genetically altered mice will elucidate the importance of TLR and ROS signaling in mediating these protective effects. Necrotizing enterocolitis (NEC) is a leading cause of morbidity and mortality in the neonatal intensive care unit. Recent laboratory and clinical studies suggest that NEC may be prevented by probiotics, which may normalize bacterial populations within the gut of premature infants and also promote cytoprotective intestinal responses. In this proposal, we plan to investigate how the probiotic commensal Lactobacillus rhamnosus reduces apoptosis, promotes proliferation, and inhibits excessive inflammation in the developing intestine of the premature infant utilizing a murine model of immature intestines.